Basic Neuroscience
High resolution 3 T fMRI in anesthetized monkeys

https://doi.org/10.1016/j.jneumeth.2011.12.023Get rights and content

Abstract

Although there are numerous 3 T MRI research devices all over the world, only a few functional studies at 3 T have been done in anesthetized monkeys. In the past, anesthetized preparations were reported to be misleading when exploring cortical brain regions outside the primary sensory areas. Nonetheless, a great improvement has been achieved in the limited effect of anesthetic agents on the reactivity of the brain.

Here, we re-address the feasibility and potential applications of the brain oxygen level dependent (BOLD) fMRI signal in Macaca mulatta monkeys that have been lightly anesthetized with sevoflurane and curarized. The monkeys were studied with commercially available coils and sequences using a 3 T clinical magnet. We obtained sagittal T1 scout images, gray matter double inversion recovery, standard gradient echo sequences and gradient echo functional imaging sequences. Given that fMRI signals are most readily identified in the cerebral cortices, we optimized Echo Planar Imaging sequences to reproduce significant changes in the BOLD signal subsequent to a visual stimulation paradigm.

Our results provide a satisfactory signal to noise ratio with a limited standard deviation range, when compared with studies on alert macaques.

We suggest that the 3 T magnet remains a valuable tool to analyze neural pathways in the macaque brain under light anesthesia and report the use of spatially resolved fMRI in higher visual areas of anesthetized monkeys. This methodology avoids the need for time-consuming training of awake monkeys, is stable over many hours, provides reproducible data and could be applied successfully to future functional studies.

Highlights

► We performed a fMRI study with anesthetized Macaca mulatta monkeys at 3 T. ► Evidence of pathway functionality of the visual system in a non-behaving primate. ► A complete overview has been provided for functional imaging of pathways. ► Opens the possibility of 3 T fMRI with anesthetized primates to other complex attempts.

Introduction

Magnetic resonance imaging (MRI) techniques are widely used and contribute to the understanding of the brain in health and disease. This methodology allows the examination of the global properties of the brain in a non-invasive way.

Neuronal activation is accompanied by an increase in both energy metabolism and local cerebral blood flow (Kida and Hyder, 2006). These physiological functions permit the identification of brain activation through the use of blood oxygenation level-dependent (BOLD) contrast in functional MRI (fMRI). BOLD-fMRI is sensitive to the decrease in deoxyhemoglobin concentration during neuronal activation (Ogawa et al., 1990). On the basis of this unique property, studies with fMRI have multiplied in human and non-human primates (NHP).

fMRI studies can be conducted in animal experimental protocols, with behaving or with anesthetized preparations. Work has been performed with behaving macaque monkeys in functional studies (Goense et al., 2010) but, a major constraint on such procedures is the fact that only a very limited number of macaques per protocol can be trained and are finally used for fMRI studies (Goense et al., 2010, Joseph et al., 2006). In fact, this is the principal advantage of the use of anesthetic preparations in functional studies, but it is thought that anesthesia would greatly influence, or even invalidate, the results obtained in both basic electrophysiological and MRI investigations (Sloan and Erian, 1993). While this concept may have been relevant in the past (see below), today's generation of anesthetic agents appear more suited to studies of the cerebral circulation if used at an appropriate concentration (Ishizawa, 2007).

The effects of anesthesia on the cerebral circulation and metabolism have always been of concern to physiologists. It has been well documented in humans that barbiturates depress both cerebral blood flow (CBF) and metabolism (CMRO2) (Kida and Hyder, 2006), and that halogenic agents such as halothane and isoflurane can alter the reactivity of the cerebral circulation to an external challenge by, for example, hypo- or hypercapnia (Goode et al., 2009). However, the more recent generation of halogenic agents, which includes desflurane and sevoflurane, when used at concentrations that do not induce surgical anesthesia, may have a positive role to play in study of the functional reactivity of the brain to a stimulus that is repeated over time. In addition, the use of curare allows muscle relaxation without affecting the neuromuscular junction, so the neuronal circuits are preserved (Sloan and Erian, 1993).

Indeed, if it could be demonstrated that a light maintenance anesthetic regime could be employed during functional activation of the brain, and that fMRI could be used to map changes in the resulting BOLD signal, then such a methodology could be used to circumvent the problems associated with the use of awake primates (environment habituation, control of physiological and biochemical parameters, movement, experimental studies of long duration) through the use of a readily available clinical 3 T camera.

Therefore, we have re-addressed the use of anesthetized NHP in fMRI studies following a visual stimulation paradigm. To circumvent the problems associated with the use of awake primates, we have addressed the feasibility of fMRI monitoring of the BOLD signal in lightly anesthetized and curarized monkeys (Macaca mulatta) with a clinical 3 T camera.

Given that activity in primary cortical structures is readily recognizable from the fMRI signal, we optimized Echo Planar Imaging (EPI) sequences to identify the BOLD signal following a visual stimulation paradigm, as already published by other authors in awake monkeys. We show that the BOLD response to the visual stimulation is strongly repeatable in anesthetized monkeys and that changes in intensity are comparable with those of awake primates, as reported by other authors. Beyond reporting the feasibility of previously described spatially resolved fMRI in higher visual areas of the NHP in anesthetized M. mulatta, the present results may be of relevance to future functional studies of the central nervous system in anesthetized primates with available clinical 3 T magnets.

Section snippets

Methods

The study was conducted on two adult male rhesus monkeys (M. mulatta, 0303 and 0390, weighing 6 and 7 kg, respectively). All experiments were performed during daytime. A veterinarian skilled in the healthcare and maintenance of NHP supervised all aspects of animal care. Animals were checked at least daily by a competent person. These checks ensure that all sick or injured animals are identified and appropriate action is taken.

The experimental procedures were performed in accordance with the

Results

We obtained a BOLD activity signal in the monkeys anesthetized with 1.0–1.5% sevoflurane after binocular visual stimuli under the different conditions that were tested. The signal modulations over time of the baseline period preceding the onset of the stimulus are presented in Table 2 for the visual cortex region of interest: mean of signal modulations over time, the mean standard deviation over time, the signal to noise ratio (mean intensity signal over time divided by the standard deviation

Discussion

We have shown that the anesthetized M. mulatta brain can readily activate functional pathways that are identified robustly by commercially available magnets, coils and sequences with 3 T fMRI. We have provided evidence of pathway functionality in the visual system, the simplest pathways to stimulate using a MRI device. Beyond reporting the feasibility of spatially resolved fMRI in higher primary visual areas of M. mulatta, the present results may be of relevance to future functional studies with

Conclusions

The anesthetized macaque brain can readily activate functional pathways that are robustly identified by commercially available 3 T fMRI. We provide methodological evidence of functional pathways in the visual system, the simplest way to quantify activation in an MRI. We have used commercially made coils available for human clinical MRI studies in an effort to get nearer clinical research in humans and NHP. The general interest of this paper is that it opens up the possibility of fMRI 3 T studies

Acknowledgements

The authors wish to thank Tho Hai Nguyen and Laurent Chazalviel for their technical support. We also wish gratefully to thank Nicolas Delcroix for his availability and his great help with statistical analysis.

This study was supported by the French ANR MNP 2008 DyProBag program.

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